Plasmacytoid dendritic cells (pDCs) are distinct immune cells that rapidly secrete massive amounts of type I interferon (interferon ?/?, IFN) in response to nucleic acids that enter the endosomal compartment. Virus-induced IFN production by pDCs is critical for the control of viral infections, whereas aberrant pDC activation is associated with autoimmune diseases. Thus, pDCs represent a key innate immune lineage and an important target for immunotherapy. The pDCs appear related to conventional dendritic cells (cDCs) yet exhibit many properties of lymphocytes; hence the molecular basis of pDC development and lineage identity has been poorly understood. In the first cycle of the award, we have identified E protein transcription factor E2-2 as an essential and specific regulator of pDC development and maintenance. We proposed a model whereby pDCs develop in a common pathway with cDCs but get diverted into a distinct lymphocyte-like state by the induction of E2-2 and repression of its inhibitor Id2. We will now characterize the specificity, mechanism and functional targets of E2-2 activity in the pDC lineage. In Aim 1, we will test the function of a pDC-specific E2-2 isoform and simultaneously visualize E2-2 and Id2 expression in single cells during pDC development. In Aim 2, we will characterize a transcriptional co-repressor that may function together with E2-2 to inhibit Id2 expression and promote pDC development and functionality. In Aim 3, we will characterize a novel pDC-specific transcription factor that contros the migration and homeostasis of peripheral pDCs. Collectively, these studies would further elucidate the unique lineage identity of pDCs, and characterize the molecular basis of pDC development and function in the steady state, antiviral immune responses and autoimmunity.